4 research outputs found
A Second Look or, Not to Mention the Occasional Capsizing of a Windsurfer
Of
all of the epithelial ovarian cancers (EOC), clear cell adenocarcinoma
(CCA) has the worst clinical prognosis. Furthermore, the conventional
EOC biomarker CA125 is more often negative in CCA than in other subtypes
of EOC. This study sought to discover a new diagnostic biomarker that
would allow more reliable detection of CCA. Using mass spectrometry,
we compared proteins in conditioned media from cell lines derived
from CCA and other types of EOC. We identified 30 extracellular or
released proteins specifically present in CCA-derived cell lines.
Bioinformatics analyses identified a serine protease inhibitor, tissue
factor pathway inhibitor 2 (TFPI2), as a potential biomarker for CCA.
Real time RT-PCR and Western blot analyses revealed that TFPI2 was
exclusively expressed in CCA-derived cell lines and tissues. For clinical
validation, we measured levels of TFPI2 and CA125 in a set of sera
from 30 healthy women, 30 patients with endometriosis, and 50 patients
with CCA, using an automated enzyme-linked immunosorbent assay systems.
Serum levels of TFPI2 were significantly elevated in CCA patients,
even those with normal CA125 levels. In terms of area under the receiver
operating characteristic curve (AUC), TFPI2 was superior to CA125
in discriminating CCA patients from healthy women (AUC 0.97 for TFPI2
versus AUC 0.80 for CA125), or from patients with endometriosis (AUC
0.93 for TFPI2 versus 0.80 for CA125). This is the first evidence
for TFPI2 as a serum biomarker of CCA. We propose that this biomarker
may be useful for detection of CCA and for monitoring the transformation
from endometriosis into CCA
Mass Spectrometric Identification of Glycosylphosphatidylinositol-Anchored Peptides
Glycosylphosphatidylinositol
(GPI) anchoring is a post-translational
modification widely observed among eukaryotic membrane proteins. GPI
anchors are attached to proteins via the carboxy-terminus in the outer
leaflet of the cell membrane, where GPI-anchored proteins (GPI-APs)
perform important functions as coreceptors and enzymes. Precursors
of GPI-APs (Pre-GPI-APs) contain a C-terminal hydrophobic sequence
that is involved in cleavage of the signal sequence from the protein
and addition of the GPI anchor by the transamidase complex. In order
to confirm that a given protein contains a GPI anchor, it is essential
to identify the C-terminal peptide containing the GPI-anchor modification
site (ω-site). Previously, efficient identification of GPI-anchored
C-terminal peptides by mass spectrometry has been difficult, in part
because of complex structure of the GPI-anchor moiety. We developed
a method to experimentally identify GPI-APs and their ω-sites.
In this method, a part of GPI-anchor moieties are removed from GPI-anchored
peptides using phosphatidylinositol-specific phospholipase C (PI-PLC)
and aqueous hydrogen fluoride (HF), and peptide sequence is then determined
by mass spectrometry. Using this method, we successfully identified
10 GPI-APs and 12 ω-sites in the cultured ovarian adenocarcinoma
cells, demonstrating that this method is useful for identifying efficiently
GPI-APs
Mass Spectrometric Identification of Glycosylphosphatidylinositol-Anchored Peptides
Glycosylphosphatidylinositol
(GPI) anchoring is a post-translational
modification widely observed among eukaryotic membrane proteins. GPI
anchors are attached to proteins via the carboxy-terminus in the outer
leaflet of the cell membrane, where GPI-anchored proteins (GPI-APs)
perform important functions as coreceptors and enzymes. Precursors
of GPI-APs (Pre-GPI-APs) contain a C-terminal hydrophobic sequence
that is involved in cleavage of the signal sequence from the protein
and addition of the GPI anchor by the transamidase complex. In order
to confirm that a given protein contains a GPI anchor, it is essential
to identify the C-terminal peptide containing the GPI-anchor modification
site (ω-site). Previously, efficient identification of GPI-anchored
C-terminal peptides by mass spectrometry has been difficult, in part
because of complex structure of the GPI-anchor moiety. We developed
a method to experimentally identify GPI-APs and their ω-sites.
In this method, a part of GPI-anchor moieties are removed from GPI-anchored
peptides using phosphatidylinositol-specific phospholipase C (PI-PLC)
and aqueous hydrogen fluoride (HF), and peptide sequence is then determined
by mass spectrometry. Using this method, we successfully identified
10 GPI-APs and 12 ω-sites in the cultured ovarian adenocarcinoma
cells, demonstrating that this method is useful for identifying efficiently
GPI-APs
Secretome-Based Identification of TFPI2, A Novel Serum Biomarker for Detection of Ovarian Clear Cell Adenocarcinoma
Of
all of the epithelial ovarian cancers (EOC), clear cell adenocarcinoma
(CCA) has the worst clinical prognosis. Furthermore, the conventional
EOC biomarker CA125 is more often negative in CCA than in other subtypes
of EOC. This study sought to discover a new diagnostic biomarker that
would allow more reliable detection of CCA. Using mass spectrometry,
we compared proteins in conditioned media from cell lines derived
from CCA and other types of EOC. We identified 30 extracellular or
released proteins specifically present in CCA-derived cell lines.
Bioinformatics analyses identified a serine protease inhibitor, tissue
factor pathway inhibitor 2 (TFPI2), as a potential biomarker for CCA.
Real time RT-PCR and Western blot analyses revealed that TFPI2 was
exclusively expressed in CCA-derived cell lines and tissues. For clinical
validation, we measured levels of TFPI2 and CA125 in a set of sera
from 30 healthy women, 30 patients with endometriosis, and 50 patients
with CCA, using an automated enzyme-linked immunosorbent assay systems.
Serum levels of TFPI2 were significantly elevated in CCA patients,
even those with normal CA125 levels. In terms of area under the receiver
operating characteristic curve (AUC), TFPI2 was superior to CA125
in discriminating CCA patients from healthy women (AUC 0.97 for TFPI2
versus AUC 0.80 for CA125), or from patients with endometriosis (AUC
0.93 for TFPI2 versus 0.80 for CA125). This is the first evidence
for TFPI2 as a serum biomarker of CCA. We propose that this biomarker
may be useful for detection of CCA and for monitoring the transformation
from endometriosis into CCA